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Abstract

BeetleBase (http://www.beetlebase.org) has been updated to provide more comprehensive genomic information for the red flour beetle Tribolium castaneum. The database contains genomic sequence scaffolds mapped to 10 linkage groups (genome assembly release Tcas_3.0), genetic linkage maps, the official gene set, Reference Sequences from NCBI (RefSeq), predicted gene models, ESTs and whole-genome tiling array data representing several developmental stages. The database was reconstructed using the upgraded Generic Model Organism Database (GMOD) modules. The genomic data is stored in a PostgreSQL relatational database using the Chado schema and visualized as tracks in GBrowse. The updated genetic map is visualized using the comparative genetic map viewer CMAP. To enhance the database search capabilities, the BLAST and BLAT search tools have been integrated with the GMOD tools. BeetleBase serves as a long-term repository for Tribolium genomic data, and is compatible with other model organism databases.

INTRODUCTION

The red flour beetle, Tribolium castaneum, is a sophisticated genetic model organism for studies of insect development and pest biology as well as comparative genomics (1,2). A burgeoning wealth of genomic information, including a whole genome shotgun (WGS) assembly of the genome sequence, has become available in recent years (2).

BeetleBase was developed to provide a centralized database to serve the growing needs of the Tribolium research community. The first version of BeetleBase (3) was based on genome release Tcas_1.0. The database provided access to unmapped scaffolds, Fgenesh predicted genes, genetic markers, BAC-end sequences and ESTs. The database was implemented using an early version of the Chado schema, GBrowse, and CMAP developed by the Generic Model Organism Database (GMOD) project (http://www.gmod.org/).

Additional data has become available since the initial release of BeetleBase, including updates to the genome assembly and annotation, necessitating updates to the BeetleBase schema and data content. Here we present an updated version of BeetleBase (http://www.beetlebase.org), implemented using recent versions of Generic Model Organism Database (GMOD) tools, and integrated with BLAST (4) and BLAT (5) alignment tools. Data content has been expanded to include the latest version of the genome assembly (Tcas_3.0), a comprehensive collection of gene sets including the first unified release of the Tribolium Official Gene Set (OGS), EST alignments to the genome, and whole-genome transcriptional information from DNA tiling array experiments. These updates will provide a valuable resource for the Tribolium community, and serve as a foundation for integration of future datasets.

Gene models

Several gene prediction programs were previously used to annotate the Tcas_2.0 assembly, and were combined into a consensus GLEAN gene set (2,7). More than 2000 genes were manually curated by members of the Tribolium Genome Sequencing Consortium (2); however, new and updated gene models were not combined into a unified gene set. We generated the first Tribolium Official Gene Set (OGS) by merging the GLEAN gene set with the manually curated gene annotations. First, each manually curated gene was mapped to the Tcas_3.0 assembly, and the validity of the mRNA, CDS, and/or peptide sequence for each of the manually curated genes was checked to ensure that the peptide sequence represented the same exons/splice sites as the mRNA and CDS sequences. Incorrectly annotated exons were replaced with exon coordinates determined by Splign (8). In some cases, genes required manual curation to determine the correct or most likely gene structure. A non-redundant Official Gene Set was constructed by merging the GLEAN and manually curated gene sets, automatically replacing GLEAN models with overlapping, manually curated models. Finally, unique identifiers were assigned to each gene to facilitate communication of Tribolium genes in research publications. A total of 16 561 official genes were generated, assigned identifiers such as ‘TC######’, and submitted to NCBI (Table 2).

Since some researchers may benefit from access to other gene sets, we migrated the AUGUSTUS, Ensembl, Fgenesh, NCBI supported and ab initio gene models, and the combined GLEAN genes from the Genboree database hosted at Baylor College of Medicine (http://www.genboree.org), and converted their coordinates into the genome coordinates of Tcas_3.0 (Table 2). Only a few of the predicted genes were lost when the scaffolds were reorganized. In addition, the latest RefSeq annotation from NCBI (build 2.1, based on the same Tcas_3.0 assembly), which includes 3613 protein accessions that are new or changed from the original annotation run used for the GLEAN set, was imported into BeetleBase. Each gene set is available for viewing in a separate track in GBrowse (Figure 1).

Tribolium gene models. Various gene models are shown in different tracks for easy comparison. By clicking one of the gene models, detailed information can be retrieved. The RefSeq gene models are linked to NCBI Entrez Gene report pages. The Tribolium...

BeetleBase provides a detailed gene report page for each gene in the OGS, which can be accessed and managed through the web interface (Figure 2). The new BeetleBase database provides more comprehensive gene information than was previously available, including detailed information on the gene structure, nomenclature, and additional annotation data provided during the manual curation process. Links are also provided to overlapping RefSeq Gene and transcript records at NCBI (9,10) to facilitate use of data from both databases.

Official gene report page. The report page provides detailed information for an official gene. The information can be modified by clicking ‘Edit’ after obtaining a password from the BeetleBase webmaster. The report page links to GBrowse...

EST and BAC-end sequences

EST and BAC-end sequences were downloaded from the dbEST and GSS databases, respectively, at NCBI. 55 616 ESTs from five different tissue- and stage-specific cDNA libraries (11) were cleaned and polyA sequences removed using in-house software tools (http://bioinformatics.ksu.edu/ArthropodEST) and aligned to the genome using the Exonerate algorithm (12). A total of 50 277 EST-genome alignments were generated. BAC-end sequences from the Tribolium BAC library (constructed by Exelixis, Inc, South San Francisco, CA and archived for distribution by the Clemson University Genomics Institute (https://www.genome.clemson.edu/) were mapped to the genome and added to the database. Out of 28 788 BAC-end sequences, 27 810 were aligned to the genome using BLAST.

Tiling arrays

Using Roche NimbleGen HD2 whole genome DNA tiling arrays, whole genome expression data have been collected for several developmental stages including three embryonic, the last larval, three pupal and two adult stages. Briefly, fluorescently labeled cDNAs were hybridized to the custom-designed microarrays; GFF files were constructed using the signal intensities from each feature on the array that were quantified directly from the scanned array images without any data management such as background subtraction. This was done to provide immediate access to the data to help verify computed gene models and assist manual annotation efforts while the data are processed and further analyzed. Figure 3 shows empirically derived transcriptome tiling array data generated from several developmental stages for a portion of the Tribolium genome.

Tribolium genome tiling arrays. Tracks for 11 developmental time points are shown. Blue vertical bars of each track represent fluorescence intensity values for oligonucleotide probes tiled across a 54 kb region on the array. The official gene mRNA track...

FTP site

Datasets that can be downloaded from BeetleBase (ftp://bioinformatics.ksu.edu/pub/BeetleBase/3.0/) include the contig sequences, GFF (.gff3) and assembly files (.agp) for Tcas_3.0. Sequences of GLEAN genes, GLEAN cDNAs and GLEAN peptides as well as the corresponding files for the OGS are also available.

IMPLEMENTATION

All the genomic information was compiled into Genetic Feature Format Version 3 (GFF3), which is the most common extension of GFF. The compiled information was implemented using GMOD tools (http://www.gmod.org) such as PostgreSQL-based Chado 1.0, GBrowse 1.68, and CMAP 1.0. To query sequences against the Tribolium genome, we also installed stand alone BLAST and BLAT servers. In this release of BeetleBase, we improved the usability by integrating these components.

FUNDING

Grant number P20 RR016475 from the National Center for Research Resources (NCRR), a component of the National Institutes of Health. Work at NCBI was supported by the Intramural Research Program of the NIH, National Library of Medicine. Funding for open access charge: National Center for Research Resources National Institutes of Health (P20 RR016475).